1. Field of the Invention
The present invention relates to a promoter. More specifically, the present invention relates to a promoter capable of regulating, by means of external stimulus, the gene expression of a foreign protein that is inserted into the downstream of the promoter.
2. Description of the Related Art
It is known that nitrogen is an important nutrient for plants and thus most of plants have a nitrate-phile property, and that the proliferation of plant culture cells and the growth of a plant are greatly affected by the presence or absence of nitrate or its concentration. A group of proteins functioning when a plant absorbs nitrate from its roots and transports the absorbed nitrate from a cell to another cell in the plant are known as nitrate transporter proteins.
Nitrate transporter proteins are known to include low-affinity nitrate transporters which function at the time when the nitrate concentration is high (i.e., 0.5 mM or more) and high-affinity nitrate transporters which function at the time when the nitrate concentration is low (i.e., 0.5 mM or less). A gene encoding each of two types of the nitrate transporters (hereinafter, also referred to as a low-affinity nitrate transporter gene and a high-affinity nitrate transporter gene) is reported (Kagaku to Seibutsu (Chemistry and Biology), Vol. 38, pp. 196–203, 2000). As a result of the progress of the study on molecular biology so far, the low-affinity nitrate transporter genes are reported to include CHL1 (AtNRT1) derived from Arabidopsis thaliana (Cell, Vol. 72, pp. 705–713, 1993; The Plant Cell, Vol. 8, pp. 2183–2191, 1996); NTL1 (AtNRT1:2) derived from Arabidopsis thaliana (The Plant Cell, Vol. 11, pp. 1381–1392, 1999); OsNRT1 derived from rice (Plant Physiol., Vol. 122, pp. 379–388, 2000); BnNRT1:2 derived from rapeseed (J. Biol. Chem., Vol. 273, pp. 1201, 1998); and LeNRT1 derived from tomato (Proc. Natl. Acad. Sci. USA, Vol. 93, pp. 8139–8144, 1996). In addition, the CHL1 is also reported to have both functions of low and high affinities (The Plant Cell, Vol. 11, pp. 865–874, 1999). Of these genes, it is experimentally confirmed that CHL1, NTL1 and OsNRT1 have the nitrate transport activity. Besides these genes, although many homologous genes are isolated, they are not determined for nitrate transport activity, and thus they are not shown to be nitrate transporter genes in the present circumstances.
As described above, nitrate transporters have also thus far been investigated for the respective gene and the protein functions and many reports have been published; however, no promoters of nitrate transporter genes have been reported.
In general, a promoter, an expression regulatory region of the gene, is capable of regulating the expression of a gene located downstream of the promoter, and therefore the selection of a promoter to be used for regulating the expression of a foreign gene is important. In other words, the selection of the promoter is important in the preparation of a transgenic plant, because the promoter is capable of controlling the stage, tissue and intensity in the expression of the foreign gene. Methods that are frequently used to express a foreign gene in a plant cell include a method of connecting a foreign gene to the downstream of a CaMV 35S promoter, or a nopaline synthase gene promoter (Sander P. R. et al., Nucleic Acid RES, 15 (1987) 1543–1558), etc. Methods of regulating, by the application of external stimulus, the expression of a foreign gene have been reported; examples include a method of making use of a promoter of the α-amylase gene (Chan M. T. et al., J. Biol. Chem., 269 (1994) 17635–17641), a promoter of the phosphate transporter gene (N. Mitsukawa et al., Proc. Natl. Acad. Sci. USA, 94 (1997) 7098–7102), or a damage-inducible promoter (T. Yamada et al., Plant cell Physiol. (1994) 917–926), a chemical-inducible promoter (Ward et al., Plant Mol. Biol., 22 (1993) 361–366), and a photoinducible promoter (Fluhr et al., Science, 232 (1986) 1106–1112). However, the presence of a promoter that induces the expression of a gene by means of nitrate stimulus has never been reported.